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pKa calculations for class A beta-lactamases: influence of substrate binding
J Lamotte-Brasseur1, V Lounnas, X Raquet
1Centre d'Ingénierie des Protéines, University of Liège, Sart-Tilman, Belgium. jlamotte@ulg.ac.be
Protein Science : a Publication of the Protein Society
|February 27, 1999
Summary
The general base in beta-lactamase catalysis is not Lys73, as its pKa remains high even with substrate binding. Lys234 likely stabilizes the tetrahedral intermediate, impacting bacterial resistance mechanisms.
Area of Science:
- Biochemistry
- Enzymology
- Molecular Biology
Background:
- Beta-lactamases confer bacterial resistance to beta-lactam antibiotics, a critical clinical challenge.
- The precise identity of the general base catalyst in beta-lactamase action remains debated, with Glu166 and Lys73 as primary candidates.
Purpose of the Study:
- To investigate the role of Lys73 as a general base in the catalytic mechanism of beta-lactamases.
- To determine the pKa of Lys73 in the presence of beta-lactam substrates within the Henri-Michaelis complex and tetrahedral intermediate.
Main Methods:
- Continuum electrostatic calculations were employed.
- pKa values of Lys73 were estimated for five wild-type and three mutant beta-lactamases.
- Calculations considered the Henri-Michaelis complex and the tetrahedral intermediate.
Main Results:
- The computed pKa of Lys73 remained above 10 in all tested conditions, even with bound substrate.
- This indicates Lys73 is unlikely to function as a proton abstractor.
- The pKa of Lys234 increased in the tetrahedral intermediate, suggesting its role in stabilization.
Conclusions:
- Lys73 is unlikely to be the general base catalyst in beta-lactamase activity.
- Lys234 likely plays a role in stabilizing the tetrahedral intermediate.
- The study provides insights into the catalytic mechanisms of beta-lactamases and bacterial resistance.